In vitro propagation of Paphiopedilum orchid through formation of protocorm-like bodies

Abstract

Paphiopedilum orchids are among the world's most popular orchid due to their impressively beautiful flowers. Propagation of these orchid genera has been hampered by the naturally slow growth rate of the plant, which renders it very difficult to be propagated through conventional methods. In vitro culture techniques have provided a useful alternative technology for propagating this recalcitrant species. In this study, the propagation of P. rothschildianum was achieved through the in vitro formation of secondary protocorm-like bodies (PLBs) from the primary PLB that developed from stem-derived callus. The PLBs were cultured on half-strength MS medium supplemented with different concentrations (1.0, 2.0, 3.0, and 4.0 lM) of 6-benzyladenine (BA) and kinetin for the induction of secondary PLBs. The highest number of secondary PLBs formed was obtained on half-strength MS medium supplemented with 4.0 lM kinetin, with an average of 4.1 PLBs per explant after 8 weeks of culture. The secondary PLBs continued to proliferate further and formed 9.5-12.1 new PLBs per secondary PLB after being subcultured onto half-strength plant growth regulator-free MS medium supplemented with 60 g/L banana homogenate (BH). These tertiary PLBs were subcultured onto media containing different organic additives, such as BH, coconut water, potato homogenate, and tomato homogenate, for plantlet regeneration. Among the organic additives tested, the addition of 20% CW to half-strength MS medium resulted in the best average plantlet regeneration percentage from the PLBs, 67.9%, after 8 weeks of culture.

Genetic Stability of In vitro Multiplied Phalaenopsis gigantea Protocorm-like Bodies as Affected by Chitosan

“…It has been previously reported that the presence of 20% CW in tissue culture media were effective for PLBs growth and plantlet regeneration (Ng and Saleh, 2010). This natural compound is rich in cytokinins that are mostly used for PLB multiplication in the plant tissue culture…”

Ultrastructure study of Vanda Kasem's Delight orchid's protocorm-like body

“…Interestingly, SEM analysis of anterior meristematic pole of PLB showed elongated cone-shaped meristematic dome (Figure 2a and b). A more intense meristematic activity at one side of the dome displaced the apical end forming a hook-like structure depicting differentiated cells that are regarded as orchid embryos (Ng & Saleh, 2011) which develop with two discrete bipolar structures, where the upper region becomes the shoot while the bottom region becomes the root meristem (Jones, 2009). A notable anterior pole and posterior pole was observed on the PLB (Figure 1a).…”

“…The bipolar PLB eventually differentiated anterior and posterior regions into shoot and root system respectively (Figure 1d). Thus, meristematic PLB can be induced to regenerate into whole plantlets on plant growth regulator (PGR)-free medium (Ng & Saleh, 2011). Furthermore, the shoots develop directly from the primary and secondary meristems of PLB without an intervening explantorganogenesis phase (Ng & Saleh, 2011).…”

“…Thus, meristematic PLB can be induced to regenerate into whole plantlets on plant growth regulator (PGR)-free medium (Ng & Saleh, 2011). Furthermore, the shoots develop directly from the primary and secondary meristems of PLB without an intervening explantorganogenesis phase (Ng & Saleh, 2011). Thus, absence of phytohormones and explant-organogenesis phase reduce the opportunity for somaclonal variation.…”

Mapping of the Interaction Between Agrobacterium tumefaciens and Vanda Kasem’s Delight Orchid Protocorm-Like Bodies

“…This is partly due to their ability to proliferate rapidly [4]. Furthermore, PLBs can regenerate into complete plantlets in plant growth regulator free medium due to their bipolar nature which develop into shoot and root meristems [5].…”